This project focuses on the regulation and function of the MAPK Kinase Kinase MEKK3. MAPK pathways have been shown to be essential for cell growth, differentiation and apoptosis. It is therefore of interest to gain a better molecular understanding i) of all of the component parts that make up MAPK signaling paths, and ii) of the physiologic targets of individual MAPK pathways in normal and diseased states. We originally cloned MEKK3 and determined that it can activate all four major MAPKs, namely ERK, JNK, p38 and ERK5, albeit to different extents. We also discovered that MEKK3, when overexpressed in an active form, is able to arrest growth, protect from some apoptotic insults and can activate NF-kappaB. Recently, MEKK3 was proposed to have a direct role in activating the IKK kinase complex, a critical complex that phosphorylates and thus causes degradation of the inhibitors of NF-kappaB. We have cloned an adaptor protein that directly links MEKK3 with the IKK kinase complex and we have now determined that this adaptor when overexpressed, negatively regulates TNF-mediated activation of select MAPKs. We have furthermore discovered that MEKK3 can also interact with AKT, a kinase that has been implicated in many survival pathways and which is able to activate MAPK pathways as well. These data suggest the existence of a large signaling complex in which both the NF-kappaB-activating IKK core complex as well as MAPK-activating AKT and MEKK3 kinases may reside, possibly for the purpose of coordinating the activation of several targets in response to specific signal. Evidence for other, distinct large complexes allowing for regulation of both MAPK and NF-kappaB also emerge from work with the IKK-associated CIKS adaptor, which is a strong activator of select MAPKs as well.